Type-1 polarized DCs (aDC1s) show selectively enhanced ability to induce functional Th1 and CTL (Teff) cells, when compared to the current standard of clinically-used DCs (sDC). We are currently implementing a phase l/ll clinical trial evaluating the relative abilities of DDC1 and sDCs to induce melanoma-specific immunity in stage III/IV melanoma patients (UPCI 03-118; funded by an independent R21 grant to Kalinski/Kirkwood). Our new preliminary data demonstrate that otDC1 and sDCs induce different sets of T cell-associated chemokine receptors (CKRs), with aDC1s being superior in inducing CXCR3 andCCRS, the CKRs implicated in tumor-entry of melanoma-specific T cells. We propose to analyze the mechanism of the differential ability of aDC1 and sDCs to induce distinct CRK expression in tumor-specific T cells, as the fourth DC-related signal essential for the efficacy of DC-based cancer vaccines. We will test the hypothesis type 2 responses (delivery of signal 3), can also differentially regulate the expression of CKRs in tumorthat DCs maturing in different environments, in addition to their differential ability to induce type-1 versus specific immune cells (delivery of signal 4). We will pursue three Specific Aims: Specific Aim 1. Determine the mechanism of DC-dependent regulation of T cell chemokine responsiveness in vitro. We will test the hypothesis that, in contrast to the current paradigm, the induction of a distinct CKR profile is regulated in a different mechanism than the Th1/Th2 commitment. Specific Aim 2. Demonstrate that DCs maturing in different conditions (ccDC1 v. sDC) differentially regulate chemokine responsiveness of melanoma-specific T cells in vitro and in vivo. We will directly test the hypothesis that ocDC1 and sDCs can (differentially) modulate the CKR patterns in melanoma-specific T cells, and can revert the established CKR patterns on the tumor-induced T cells in melanoma patients. Specific Aim 3. Demonstrate the differences in local chemokine production and the character of local T cell infiltrates between DTH sites, primary, and metastatic tumor lesions. We will test the hypothesis that the tumor tissues constitute a biased chemokine environment and the hypothesis that primary and metastatic tumors represent different chemokine environments, contributing to often ineffective immune control of tumor metastases, even in DTH-positive patients. The proposed studies will help us to better understand the mechanisms and paradigms of DC-dependent regulation of cancer immunity, and to prospectively develop further improved immunotherapies of melanoma and other tumors, by helping to direct the vaccination-induced tumor-specific T cells to the tumors.